The two-dimensional (2D)13C-13C spin diffusion spectra were recorded using a pulse sequence composed of a 60-kHz1H excitation 90 pulse, a linearly ramped1H-13C cross-polarization period, a 10-kHz rf-assisted diffusion1H field during the 10-ms mixing period, and a 100-kHz1H two-pulse phase modulation decoupling

The two-dimensional (2D)13C-13C spin diffusion spectra were recorded using a pulse sequence composed of a 60-kHz1H excitation 90 pulse, a linearly ramped1H-13C cross-polarization period, a 10-kHz rf-assisted diffusion1H field during the 10-ms mixing period, and a 100-kHz1H two-pulse phase modulation decoupling. fibril, phosphorylation, solid state NMR, morphology, structure == Introduction == Amyloid fibrils are -sheet-enriched fibrillar aggregates with misfolded polypeptides and proteins. The formation and deposition of these fibrils are related to a variety of neurodegenerative diseases, including Alzheimer’s disease (AD), 3Parkinson’s disease, and Huntington disease among others (14). Amyloid fibrils derived from the same primary sequences of polypeptides or proteins usually show distinct morphologiesin vitro(5, 6), and the morphologies of fibrils are sensitive to a variety of fibrillation conditions such as temperature (7), agitation (8), salt concentrations (9), surfactant (10), and seeding effects (11). Recent evidence revealed that amyloids can spread through a prion-like mechanism where fibrils seem to play important roles (1214). Different fibrils with their specific morphologies, reminiscent of prion-like strains, may cause distinct pathological phenotypes and link different structures to the variations in disease transmission and pathology (1517). Furthermore, fragmentation of fibrils, Imiquimod (Aldara) which always produces new ends for self- or cross-seeded fibrillation, is of critical importance for infectious amyloids (18, 19). Senile plaques consisting of fibrillar A are considered one of the important hallmarks in AD (20). The 40-residue and 42-residue A peptides (i. e. A40 and A42, respectively) are the two main fibrillar species. Recently, A has also been reported to exhibit prion-like propagation properties (21, 22). Distinct strains of A were discerned in Alzheimer’s patients (2326). Different amyloid propagation properties and structural profiles of A40 and A42 Imiquimod (Aldara) mimic distinct amyloid strains (10, 27). The phenotypes induced by exogenous injection of A-containing brain extracts from Alzheimer’s patients were dependent on both the hosts and the sources of Imiquimod (Aldara) agents, suggesting that polymorphic A strains might result in varying biological activities (25, 28, 29). Molecular structures of A fibrils derived from Alzheimer’s patients with distinct clinical histories were also different (24). This underlines that the structural variations of A fibrils may correlate with the variations of pathological phenotypes (24). In addition , compared with the brain-derived A HMGIC fibrils, the synthetic fibrils showed lower prion activities and different molecular structures (23, 24, 30), implying that some crucial factorsin vivomight account for different fibrillar polymorphisms and pathological phenotypes. Recently, post-translational modifications of A, such as phosphorylation and pyroglutamation, occurringin vivohave been found to promote the progression of AD (31, 32). Among different types of post-translational modifications, the phosphorylation of proteins plays crucial roles in protein folding (33). Phosphorylation can alter the structures of a protein and modulate its activities (33). Using Trp-cage as a model protein, Kardoset al. (34) reported that phosphorylation could serve as a conformational switch to trigger the transition from native to amyloid state. Significantly, we and other groups have demonstrated that phosphorylation is involved in the formation of low barrier hydrogen bond (35) and turn conformations (36) as well as the destabilization of -hairpin structure (37). Furthermore, we have also reported that phosphorylation may modulate the fibrillation process of amyloid proteins, such as Tau and -synuclein (3840). Herein, we describe a novel regulatory function of phosphorylation at Ser8on morphology, biophysical properties, cellular toxicity, and structures of the A40 fibrils. It has been shown that phosphorylation at Ser8in A has important roles in late onset Imiquimod (Aldara) sporadic AD (31, 41, 42). Phosphorylation at Ser8was found in the brains of Alzheimer’s patients in a hierarchical sequence and was specially suggested to be associated with symptomatic AD (43). Phosphorylation at Ser8was modulated by protein kinase A (44). Additionally , this site-specific phosphorylation was known to accelerate the nucleation-dependent fibrillation of A and to enhance the A-mediated amyloid toxicity (44). Attenuation of A clearance via insulin-degrading enzyme and angiotensin-converting enzyme induced by this phosphorylation was also reported (45). Furthermore, the phosphorylation at Ser8could elevate numbers of strong hydrogen bonds in the N terminus of A and increase the stability of the resulting pathogenic fibrils. The latter represents one of the crucial factors for disease progression in the brain (46). Despite all the functional importance, it is not clear whether this residue-specific phosphorylation can modify the morphologies and structures of A Imiquimod (Aldara) fibrils, which are closely related to transmission and progression.